Morpholine derivatives as antagonists of orexin receptors

ABSTRACT

This invention relates to morpholine derivatives of formula (I) and their use as antagonists of orexin receptors.

[0001] This invention relates to morpholine derivatives and their use aspharmaceuticals.

[0002] Many medically significant biological processes are mediated byproteins participating in signal transduction pathways that involveG-proteins and/or second messengers.

[0003] Polypeptides and polynucleotides encoding the human7-transmembrane G-protein coupled neuropeptide receptor, orexin-1(HFGAN72), have been identified and are disclosed in EP-A-875565,EP-A-875566 and WO 96/34877. Polypeptides and polynucleotides encoding asecond human orexin receptor, orexin-2 (HFGANP), have been identifiedand are disclosed in EP-A-893498.

[0004] Polypeptides and polynucleotides encoding polypeptides which areligands for the orexin-1 receptor, e.g. orexin-A (Lig72A) are disclosedin EP-A-849361.

[0005] Orexin receptors are found in the mammalian host and may beresponsible for many biological functions, including pathologiesincluding, but not limited to, depression; anxiety; addictions;obsessive compulsive disorder; affective neurosis/disorder; depressiveneurosis/disorder; anxiety neurosis; dysthymic disorder; behaviourdisorder; mood disorder; sexual dysfunction; psychosexual dysfunction;sex disorder; sexual disorder; schizophrenia; manic depression;delerium; dementia; severe mental retardation and dyskinesias such asHuntington's disease and Gilles de la Tourett's syndrome; disturbedbiological and circadian rhythms; feeding disorders, such as anorexia,bulimia, cachexia, and obesity; diabetes; appetite/taste disorders;vomiting/nausea; asthma; cancer; Parkinson's disease; Cushing'ssyndrome/disease; basophil adenoma; prolactinoma; hyperprolactinemia;hypopituitarism; hypophysis tumor/adenoma; hypothalamic diseases;Froehlich's syndrome; adrenohypophysis disease; hypophysis disease;hypophysis tumor/adenoma; pituitary growth hormone; adrenohypophysishypofunction; adrenohypophysis hyperfunction; hypothalamic hypogonadism;Kallman's syndrome (anosmia, hyposmia); functional or psychogenicamenorrhea; hypopituitarism; hypothalamic hypothyroidism;hypothalamic-adrenal dysfunction; idiopathic hyperprolactinemia;hypothalamic disorders of growth hormone deficiency; idiopathic growthhormone deficiency; dwarfism; gigantism; acromegaly; disturbedbiological and circadian rhythms; and sleep disturbances associated withsuch diseases as neurological disorders, neuropathic pain and restlessleg syndrome, heart and lung diseases; acute and congestive heartfailure; hypotension; hypertension; urinary retention; osteoporosis;angina pectoris; myocardial infarction; ischaemic or haemorrhagicstroke; subarachnoid haemorrhage; head injury such as sub-arachnoidhaemorrhage associated with traumatic head injury; ulcers; allergies;benign prostatic hypertrophy; chronic renal failure; renal disease;impaired glucose tolerance; migraine; hyperalgesia; pain; enhanced orexaggerated sensitivity to pain, such as hyperalgesia, causalgia andallodynia; acute pain; bum pain; atypical facial pain; neuropathic pain;back pain; complex regional pain syndromes I and II; arthritic pain;sports injury pain; pain related to infection, e.g. HIV, post-poliosyndrome, and post-herpetic neuralgia; phantom limb pain; labour pain;cancer pain; post-chemotherapy pain; post-stroke pain; post-operativepain; neuralgia; conditions associated with visceral pain includingirritable bowel syndrome, migraine and angina; urinary bladderincontinence e.g. urge incontinence; tolerance to narcotics orwithdrawal from narcotics; sleep disorders; sleep apnea; narcolepsy;insomnia; parasomnia; jet-lag syndrome; and neurodegenerative disorders,which includes nosological entities such asdisinhibition-dementia-parkinsonism-amyotrophy complex;pallido-ponto-nigral degeneration, epilepsy, and seizure disorders.

[0006] Experiments have shown that central administration of the ligandorexin-A (described in more detail below) stimulated food intake infreely-feeding rats during a 4 hour time period. This increase wasapproximately four-fold over control rats receiving vehicle. These datasuggest that orexin-A may be an endogenous regulator of appetite.Therefore, antagonists of its receptor may be useful in the treatment ofobesity and diabetes, see Cell, 1998, 92, 573-585.

[0007] There is a significant incidence of obesity in westernisedsocieties. According to WHO definitions a mean of 35% of subjects in 39studies were overweight and a further 22% clinically obese. It has beenestimated that 5.7% of all healthcare costs in the USA are a consequenceof obesity. About 85% of Type 2 diabetics are obese, and diet andexercise are of value in all diabetics. The incidence of diagnoseddiabetes in westernised countries is typically 5% and there areestimated to be an equal number undiagnosed. The incidence of bothdiseases is rising, demonstrating the inadequacy of current treatmentswhich may be either ineffective or have toxicity risks includingcardiovascular effects. Treatment of diabetes with sulfonylureas orinsulin can cause hypoglycaemia, whilst metformin causes GIside-effects. No drug treatment for Type 2 diabetes has been shown toreduce the long-term complications of the disease. Insulin sensitiserswill be useful for many diabetics, however they do not have ananti-obesity effect.

[0008] Rat sleep/EEG studies have also shown that central administrationof orexin-A, an agonist of the orexin receptors, causes a dose-relatedincrease in arousal, largely at the expense of a reduction inparadoxical sleep and slow wave sleep 2, when administered at the onsetof the normal sleep period. Therefore antagonists of its receptor may beuseful in the treatment of sleep disorders including insomnia.

[0009] The present invention provides morpholine derivatives which arenon-peptide antagonists of human orexin receptors, in particularorexin-1 receptors. In particular, these compounds are of potential usein the treatment of obesity, including obesity observed in Type 2(non-insulin-dependent) diabetes patients, and/or sleep disorders,and/or stroke, particularly ischemic or haemorrhagic stroke.

[0010] International Patent Applications WO99/09024, WO99/58533,WO00/47577, and WO00/47580, disclose phenyl urea derivatives andWO00/47576, discloses quinolinyl cinnamide derivatives as orexinreceptor antagonists.

[0011] According to the invention there is provided a compound offormula (I):

[0012] wherein:

[0013] R¹ is phenyl, naphthyl, a mono or bicyclic heteroaryl groupcontaining up to 3 heteroatoms selected from N, O and S; any of whichmay be optionally substituted;

[0014] R² represents phenyl or a 5- or 6-membered heteroaryl groupcontaining up to 3 heteroatoms selected from N, O and S, wherein thephenyl or heteroaryl group is substituted by R³, and further optionalsubstituents; or R² represents an optionally substituted bicyclicaromatic or bicyclic heteroaromatic group containing up to 3 heteroatomsselected from N, O and S;

[0015] R³ represents an optionally substituted (C₁₋₄)alkoxy, halo,optionally substituted (C₁₋₆)alkyl, optionally substituted phenyl, or anoptionally substituted 5- or 6-membered heterocyclic ring containing upto 3 heteroatoms selected from N, O and S;

[0016] or a pharmaceutically acceptable derivative thereof.

[0017] Examples of groups where R¹ is a mono or bicyclic heteroarylgroup containing up to 3 heteroatoms selected from N, O and S, includepyridyl, furanyl, indolyl, benzofuranyl, quinolinyl, isoquinolinyl,pyrazinyl, quinoxalinyl, benzoxazolyl, pyrazolyl and isoxazolyl.

[0018] Preferably R¹ is an optionally substituted phenyl, benzofuranyl,quinolinyl, indolyl or benzoxazolyl. The group may have up to 5,preferably 1, 2 or 3 optional substituents.

[0019] Examples of groups where R² represents a 5- or 6-memberedheteroaryl group containing up to 3 heteroatoms selected from N, O andS, include thiazolyl, pyrazolyl, triazolyl, pyridazyl and isoxazolyl.

[0020] Preferably when R² represents phenyl, or a 5- or 6-memberedheterocyclyl group the substituent R³ is adjacent to the point ofattachment to the amide carbonyl group.

[0021] Preferably R² represents optionally substituted thiazolyl.Particular R² groups incorporating the R³ substituent, that may bementioned are 4-(2-methyl-5-(4-fluorophenyl))thiazolyl,4-(2-methyl-5-(3-fluorophenyl))thiazolyl,4-(2-methyl-5-(2-fluorophenyl))thiazolyl, and4-(2-methyl-5-phenyl)thiazolyl, specifically4-(2-methyl-5-(4-fluorophenyl))thiazolyl.

[0022] R³ may be a trifluoromethoxy group, halo, (C₄₋₆)alkyl, optionallysubstituted phenyl or an optionally substituted 5- or 6-memberedheterocyclyl ring contains up to 3 heteroatoms selected from N, O and S.

[0023] Examples of groups where R³ is a 5- or 6-membered heterocyclylgroup containing up to 3 heteroatoms selected from N, O and S, includefuranyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl,oxadiazolyl, thiadiazolyl, pyridyl, triazolyl, triazinyl, pyridazyl,pyrimidinyl, isothiazolyl, isoxazolyl, pyrazinyl or pyrazolyl.

[0024] More preferably R³ represents trifluoromethoxy, methoxy, halo, oroptionally substituted phenyl, pyridyl, pyrazolyl or oxadiazolyl group.

[0025] Even more preferably R³ represents an optionally substitutedphenyl, e.g. 4-fluorophenyl.

[0026] Alternatively R³ represents a pyridyl group.

[0027] Optional substituents for the groups R¹ to R³ include halogen,hydroxy, oxo, cyano, nitro, (C₁₋₄)alkyl, (C₁₋₄)alkoxy, halo(C₁₋₄)alkyl,halo(C₁₋₄)alkoxy, aryl(C₁₋₄)alkoxy, (C₁₋₄)alkylthio, hydroxy(C₁₋₄)alkyl,hydroxy(C₁₋₄)alkoxy, (C₁₋₄)alkoxy(C₁₋₄)alkyl,(C₃₋₆)cycloalkyl(C₁₋₄)alkoxy, (C₁₋₄)alkanoyl, (C₁₋₄)alkoxycarbonyl,(C₁₋₄)alkylsulfonyl, (C₁₋₄)alkylsulfonyloxy,(C₁₋₄)alkylsulfonyl(C₁₋₄)alkyl, arylsulfonyl, arylsulfonyloxy,arylsulfonyl(C₁₋₄)alkyl, (C₁₋₄)alkylsulfonamido, (C₁₋₄)alkylamido,(C₁₋₄)alkylsulfonamido(C₁₋₄)alkyl, (C₁₋₄)alkylamido(C₁₋₄)alkyl,arylsulfonamido, arylcarboxamido, arylsulfonamido(C₁₋₄)alkyl,arylcarboxamido(C₁₋₄)alkyl, aroyl, aroyl(C₁₋₄)alkyl, oraryl(C₁₋₄)alkanoyl group; a group R^(a)R^(b)N-, R^(a)OCO(CH₂)_(r),R^(a)CON(R⁴)(CH₂)_(r), R^(a)R^(b)NCO(CH₂)_(r), R^(a)R^(b)NSO₂(CH₂)_(r)or R^(a)SO₂NR^(b)(CH₂)_(r) where each of R^(a) and R^(b) independentlyrepresents a hydrogen atom or a (C₁₋₄)alkyl group or where appropriateR^(a)R^(b) forms part of a (C₃₋₆)azacyloalkane or(C₃₋₆)(2-oxo)azacycloalkane ring and r represents zero or an integerfrom 1 to 4.

[0028] Preferred substituents are (C₁₋₄)alkyl, (C₁₋₄)alkoxy, either ofwhich may be optionally substituted, halogen and cyano.

[0029] In addition R¹ may be optionally substituted by a phenyl ringoptionally substituted by a halogen, cyano, or C₁₋₄alkanoyl orC₁₋₄alkylsulfonyl group; or by a 5- or 6-membered heterocyclic ring,optionally substituted by a (C₁₋₂)alkyl or R^(a)R^(b)N-group; whereinR^(a) and R^(b) are as defined above.

[0030] In the groups R¹ to R³, substituents positioned ortho to oneanother may be linked to form a fused ring, e.g. giving a group which is2,3-ethylenedioxyphenyl.

[0031] When a halogen atom is present in the compound of formula (I) itmay be fluorine, chlorine, bromine or iodine.

[0032] When the compound of formula (I) contains an alkyl group, whetheralone or forming part of a larger group, e.g. alkoxy or alkylthio, thealkyl group may be straight chain, branched or cyclic, or combinationsthereof, it is preferably methyl or ethyl.

[0033] It will be appreciated that compounds of formula (I) may exist asR or S enantiomers. The present invention includes within its scope allsuch isomers, including mixtures. Where additional chiral centres arepresent in compounds of formula (I), the present invention includeswithin its scope all possible diastereoismers, including mixturesthereof. The different isomeric forms may be separated or resolved onefrom the other by conventional methods, or any given isomer may beobtained by conventional synthetic methods or by stereospecific orasymmetric syntheses.

[0034] It will be understood that the invention includespharmaceutically acceptable derivatives of compounds of formula (I) andthat these are included within the scope of the invention.

[0035] Particular compounds according to the invention include thosementioned in the examples and their pharmaceutically acceptablederivatives.

[0036] As used herein “pharmaceutically acceptable derivative” includesany pharmaceutically acceptable salt, ester or salt of such ester of acompound of formula (I) which, upon administration to the recipient iscapable of providing (directly or indirectly) a compound of formula (I)or an active metabolic or residue thereof.

[0037] It will be appreciated that for use in medicine the salts of thecompounds of formula (I) should be pharmaceutically acceptable. Suitablepharmaceutically acceptable salts will be apparent to those skilled inthe art and include acid addition salts formed with inorganic acids e.g.hydrochloric, hydrobromic, sulphuric, nitric or phosphoric acid; andorganic acids e.g. succinic, maleic, acetic, fumaric, citric, tartaric,benzoic, p-toluenesulfonic, methanesulfonic or naphthalenesulfonic acid.Other salts e.g. oxalates, may be used, for example in the isolation ofcompounds of formula (I) and are included within the scope of thisinvention. Also included within the scope of the invention are solvatesand hydrates of compounds of formula (I).

[0038] Certain of the compounds of formula (I) may form acid additionsalts with one or more equivalents of the acid. The present inventionincludes within its scope all possible stoichiometric andnon-stoichiometric forms.

[0039] Since the compounds of formula (I) are intended for use inpharmaceutical compositions it will readily be understood that they areeach preferably provided in substantially pure form, for example atleast 60% pure, more suitably at least 75% pure and preferably at least85%, especially at least 98% pure (% are on a weight for weight basis).Impure preparations of the compounds may be used for preparing the morepure forms used in the pharmaceutical compositions.

[0040] According to a further feature of the invention there is provideda process for the preparation of compounds of formula (I) and saltsthereof. The following schemes detail synthetic routes to compounds ofthe invention.

[0041] wherein R¹ and R² are as defined for formula (I), P is aprotecting group and L¹ and L²are leaving groups.

[0042] An example of a protecting group P is optionally substitutedbenzyl. Deprotection conditions for step (iv) conveniently utilisescatalytic hydrogenolysis in an inert solvent (e.g. using palladium oncharcoal in a lower alcohol or ethyl acetate).

[0043] Examples of suitable leaving groups L¹ and L² include halogen,hydroxy, OC(═O)alkyl OC(═O)O-alkyl and OSO₂Me. Steps (iii) and (v) maybe carried out using a wide range of known acylation conditions, e.g. inan inert solvent such as dichloromethane, in the presence of a base suchas triethylamine. Alternatively these steps may be carried out when L¹or L² represents hydroxy, in which case the reaction takes place in aninert solvent such as dichloromethane in the presence of a diimidereagent such as 1-ethyl-3-(3-dimethylaminopropyl)carbodiimidehydrochloride, and an activator such as 1-hydroxybenzotriazole.

[0044] wherein R¹ and R² are as defined for formula (I), P and P¹ areamino protecting groups as described for Scheme 1 and L¹ and L² areleaving groups as described for Scheme 1.

[0045] Examples of protecting groups P and P¹ includet-butyloxycarbonyl, trifluoroacetyl, benzyloxycarbonyl and optionallysubstituted benzyl. Deprotection conditions, step (iv) and (vi), willdepend on the particular protecting group; for the groups mentionedabove these are respectively, acid (e.g. trifluoroacetic acid indichloromethane), base (e.g. potassium carbonate in a solvent such asaqueous methanol) and catalytic hydrogenolysis in an inert solvent (e.g.using palladium on charcoal in a lower alcohol or ethyl acetate). Inscheme ², protecting groups P and P1 are selected to be different.

[0046] The starting materials for use in the processes of Schemes 1 and2 are commercially available, known in the literature or can be preparedby known methods. Within the schemes above there is scope for functionalgroup interconversion.

[0047] Schemes 1 and 2 illustrate the synthesis of racemic compounds offormula (I), from (RS)-4-benzyl-5-oxomorpholine-3-carboxuylic acid whichmay be synthesisied from (DL)-serine as described in G. R. Brown, A. J.Foubister and B. Wright, J. Chem Soc. Perkin Trans. 1, 1985, 2577.Starting from (D)- or (L)-serine, synthetic methods known to thoseskilled in the art may be used to give single enantiomers of thecompounds of formula (I).

[0048] The compounds of formula (I) may be prepared singly or ascompound libraries comprising at least 2, e.g. 5 to 1000, preferably 10to 100 compounds of formula (I). Compound libraries may be prepared by acombinatorial ‘split and mix’ approach or by multiple parallel synthesisusing either solution phase or solid phase chemistry, by proceduresknown to those skilled in the art.

[0049] Thus according to a further aspect of the invention there isprovided a compound library comprising at least 2 compounds of formula(I), or pharmaceutically acceptable derivatives thereof.

[0050] Pharmaceutically acceptable salts may be prepared conventionallyby reaction with the appropriate acid or acid derivative.

[0051] The compounds of formula (I) and their pharmaceuticallyacceptable derivatives are useful for the treatment of diseases ordisorders where an antagonist of a human orexin receptor is requiredsuch as obesity and diabetes; prolactinoma; hypoprolactinemia;hypothalamic disorders of growth hormone deficiency; idiopathic growthhormone deficiency; Cushings syndrome/disease; hypothalamic-adrenaldysfunction; dwarfism; sleep disorders; sleep apnea; narcolepsy;insomnia; parasomnia; jet-lag syndrome; sleep disturbances associatedwith diseases such as neurological disorders, neuropathic pain andrestless leg syndrome; heart and lung diseases; depression; anxiety;addictions; obsessive compulsive disorder; affective neurosis/disorder;depressive neurosis/disorder; anxiety neurosis; dysthymic disorder;behaviour disorder; mood disorder; sexual dysfunction; psychosexualdysfunction; sex disorder; sexual disorder; schizophrenia; manicdepression; delerium; dementia; bulimia and hypopituitarism. Thecompounds of formula (I) or pharmaceutically acceptable derivativesthereof are also useful in the treatment of stroke, particularlyischaemic or haemorrhagic stroke.

[0052] The compounds of formula (I) and their pharmaceuticallyacceptable derivatives are particularly useful for the treatment ofobesity, including obesity associated with Type 2 diabetes, and sleepdisorders. Additionally the compounds are useful for the treatment ofstroke.

[0053] Other diseases or disorders which may be treated in accordancewith the invention include disturbed biological and circadian rhythms;adrenohypophysis disease; hypophysis disease; hypophysis tumor/adenoma;adrenohypophysis hypofunction; functional or psychogenic amenorrhea;adrenohypophysis hyperfunction; migraine; hyperalgesia; pain; enhancedor exaggerated sensitivity to pain such as hyperalgesia, causalgia andallodynia; acute pain; burn pain; atypical facial pain; neuropathicpain; back pain; complex regional pain syndromes I and II; arthriticpain; sports injury pain; pain related to infection e.g. HIV, post-poliosyndrome and post-herpetic neuralgia; phantom limb pain; labour pain;cancer pain; post-chemotherapy pain; post-stroke pain; post-operativepain; neuralgia; and tolerance to narcotics or withdrawal fromnarcotics.

[0054] The invention also provides a method of treating or preventingdiseases or disorders where an antagonist of a human orexin receptor isrequired, which comprises administering to a subject in need thereof aneffective amount of a compound of formula (I), or a pharmaceuticallyacceptable derivative thereof.

[0055] The invention also provides a compound of formula (I), or apharmaceutically acceptable derivative thereof, for use in the treatmentor prophylaxis of diseases or disorders where an antagonist of a humanorexin receptor is required.

[0056] The invention also provides the use of a compound of formula (I),or a pharmaceutically acceptable derivative thereof, in the manufactureof a medicament for the treatment or prophylaxis of diseases ordisorders where an antagonist of a human orexin receptor is required.

[0057] For use in therapy the compounds of the invention are usuallyadministered as a pharmaceutical composition. The invention alsoprovides a pharmaceutical composition comprising a compound of formula(I), or a pharmaceutically acceptable derivative thereof, and apharmaceutically acceptable carrier.

[0058] The compounds of formula (I) and their pharmaceuticallyacceptable derivatives may be administered by any convenient method,e.g. by oral, parenteral, buccal, sublingual, nasal, rectal ortransdermal administration, and the pharmaceutical compositions adaptedaccordingly.

[0059] The compounds of formula (I) and their pharmaceuticallyacceptable derivatives which are active when given orally can beformulated as liquids or solids, e.g. as syrups, suspensions, emulsions,tablets, capsules or lozenges.

[0060] A liquid formulation will generally consist of a suspension orsolution of the active ingredient in a suitable liquid carrier(s) e.g.an aqueous solvent such as water, ethanol or glycerine, or a non-aqueoussolvent, such as polyethylene glycol or an oil. The formulation may alsocontain a suspending agent, preservative, flavouring and/or colouringagent.

[0061] A composition in the form of a tablet can be prepared using anysuitable pharmaceutical carrier(s) routinely used for preparing solidformulations, such as magnesium stearate, starch, lactose, sucrose andcellulose.

[0062] A composition in the form of a capsule can be prepared usingroutine encapsulation procedures, e.g. pellets containing the activeingredient can be prepared using standard carriers and then filled intoa hard gelatin capsule; alternatively a dispersion or suspension can beprepared using any suitable pharmaceutical carrier(s), e.g. aqueousgums, celluloses, silicates or oils and the dispersion or suspensionthen filled into a soft gelatin capsule.

[0063] Typical parenteral compositions consist of a solution orsuspension of the active ingredient in a sterile aqueous carrier orparentally acceptable oil, e.g. polyethylene glycol, polyvinylpyrrolidone, lecithin, arachis oil or sesame oil. Alternatively, thesolution can be lyophilised and then reconstituted with a suitablesolvent just prior to administration.

[0064] Compositions for nasal administration may conveniently beformulated as aerosols, drops, gels and powders. Aerosol formulationstypically comprise a solution or fine suspension of the activeingredient in a pharmaceutically acceptable aqueous or non-aqueoussolvent and are usually presented in single or multidose quantities insterile form in a sealed container which can take the form of acartridge or refill for use with an atomising device. Alternatively thesealed container may be a disposable dispensing device such as a singledose nasal inhaler or an aerosol dispenser fitted with a metering valve.Where the dosage form comprises an aerosol dispenser, it will contain apropellant which can be a compressed gas e.g. air, or an organicpropellant such as a fluorochlorohydrocarbon or hydrofluorocarbon.Aerosol dosage forms can also take the form of pump-atomisers.

[0065] Compositions suitable for buccal or sublingual administrationinclude tablets, lozenges and pastilles where the active ingredient isformulated with a carrier such as sugar and acacia, tragacanth, orgelatin and glycerin.

[0066] Compositions for rectal administration are conveniently in theform of suppositories containing a conventional suppository base such ascocoa butter.

[0067] Compositions suitable for transdermal administration includeointments, gels and patches.

[0068] Preferably the composition is in unit dose form such as a tablet,capsule or ampoule.

[0069] The dose of the compound of formula (I), or a pharmaceuticallyacceptable derivative thereof, used in the treatment or prophylaxis ofthe abovementioned disorders or diseases will vary in the usual way withthe particular disorder or disease being treated, the weight of thesubject and other similar factors. However, as a general rule, suitableunit doses may be 0.05 to 1000 mg, more suitably 0.05 to 500 mg. Unitdoses may be administered more than once a day for example two or threetimes a day, so that the total daily dosage is in the range of about0.01 to 100 mg/kg; and such therapy may extend for a number of weeks ormonths. In the case of pharmaceutically acceptable derivatives the abovefigures are calculated as the parent compound of formula (I).

[0070] No toxicological effects are indicated/expected when a compoundof formula (I) is administered in the above mentioned dosage range.

[0071] Human orexin-A has the amino acid sequence: pyroGlu Pro Leu ProAsp Cys Cys Arg Gln Lys Thr  1              5                  10 CysSer Cys Arg Leu Tyr Glu Leu Leu His Gly Ala       15                       20 Gly Asn His Ala Ala Gly Ile Leu ThrLeu-NH₂     25                  30

[0072] Orexin-A can be employed in screening procedures for compoundswhich inhibit the ligand's activation of the orexin-1 receptor.

[0073] In general, such screening procedures involve providingappropriate cells which express the orexin-1 receptor on their surface.Such cells include cells from mammals, yeast, Drosophila or E. coli. Inparticular, a polynucleotide encoding the orexin-1 receptor is used totransfect cells to express the receptor. The expressed receptor is thencontacted with a test compound and an orexin-1 receptor ligand toobserve inhibition of a functional response. One such screeningprocedure involves the use of melanophores which are transfected toexpress the orexin-1 receptor, as described in WO 92/01810.

[0074] Another screening procedure involves introducing RNA encoding theorexin-1 receptor into Xenopus oocytes to transiently express thereceptor. The receptor oocytes are then contacted with a receptor ligandand a test compound, followed by detection of inhibition of a signal inthe case of screening for compounds which are thought to inhibitactivation of the receptor by the ligand.

[0075] Another method involves screening for compounds which inhibitactivation of the receptor by determining inhibition of binding of alabelled orexin-1 receptor ligand to cells which have the receptor ontheir surface. This method involves transfecting a eukaryotic cell withDNA encoding the orexin-1 receptor such that the cell expresses thereceptor on its surface and contacting the cell or cell membranepreparation with a compound in the presence of a labelled form of anorexin-1 receptor ligand. The ligand may contain a radioactive label.The amount of labelled ligand bound to the receptors is measured, e.g.by measuring radioactivity.

[0076] Yet another screening technique involves the use of FLIPRequipment for high throughput screening of test compounds that inhibitmobilisation of intracellular calcium ions, or other ions, by affectingthe interaction of an orexin-1 receptor ligand with the orexin-1receptor.

[0077] All publications, including but not limited to patents and patentapplications, cited in this specification are herein incorporated byreference as if each individual publication were specifically andindividually indicated to be incorporated by reference herein as thoughfully set forth.

[0078] The following Examples illustrate the preparation ofpharmacologically active compounds of the invention. The DescriptionsD1-D8 illustrate the preparation of intermediates to compounds of theinvention.

[0079] Abbreviation used herein are as follow:

[0080] MDC represents methylene dichloride

[0081] THF represents tetrahydrofuran

[0082] DMSO represents methyl sulphoxide

[0083] Description 1: (RS)-4-Benzyl-3-carboxamido-5-oxomorpholine

[0084] To (RS)-4-benzyl-5-oxo-morpholine-3-carboxylic acid (2.0 g, 8.51mmol) in MDC (40 ml) was slowly added oxalyl chloride (1.51 ml, 17.1mmol). The mixture was stirred for 0.5 h at room temperature and thenevaporated to give (RS)-4-benzyl-5-oxomorpholine-3-carbonyl chloride asa brown solid (1.99 g, 92%). Ammonia gas was bubbled steadily through asolution of (RS)-4-benzyl-5-oxomorpholine-3-carbonyl chloride (1.99 g,7.79 mmol) in MDC (40 ml) at 0° C. for 0.5 h, and then the mixtureallowed to warm to room temperature. The solid was collected byfiltration, washed with MDC (30 ml) and dried in vacuo at 40° C. to givethe title compound as a white solid (2.1 g, 99%). Mass spectrum (API⁺):Found 235 (MH³⁰ ). C₁₂H₁₄N₂O₃ requires 234.

[0085] Description 2: (RS)-3-Aminomethyl-4-benzylmorpholine

[0086] Lithium aluminium hydride (1M in THF) (20 ml, 20 mmol) was addedto a stirred solution of D1 (1.0 g, 4.27 mmol) in anhydrous THF (30 ml).The resultant mixture was stirred at ambient temperature for 24 hours,then water (0.76 ml), 8% NaOH (1.5 ml) and water (0.76 ml) were slowlyadded sequentially. The aqueous was extracted with diethyl ether (3×20ml) and the extracts dried (Na₂SO₄) and evaporated to give the titlecompound as a clear gum (0.79 g, 89%). Mass spectrum (API⁺): 207 (MH⁺).C₁₂H₁₈N₂O requires 206.

[0087] Description 3:(RS)-4-Benzyl-3-(2-methoxybenzamidomethyl)morpholine

[0088] To a solution of D2 (500 mg, 2.43 mmol) was added triethylamine(1 ml, 7.29 mmol) followed by 2-methoxybenzoyl chloride (455 mg, 2.67mmol). The resulting solution was stirred for 0.75 h at roomtemperature, then washed with saturated aqueous sodium hydrogencarbonate, dried (Na₂SO₄) and evaporated in vacuo. The residue waschromatographed on silica gel using 30-100% ethyl acetate in hexanegradient elution to give the title compound as a colourless oil (678 mg,82%). Mass spectrum (API⁺): 341 (MH⁺). C₂₀H₂₄N₂O₃ requires 340.

[0089] Description 4: (RS)-3-(2-Methoxybenzamidomethyl)morpholine

[0090] To a solution of D3 (1.12 g, 3.29 mmol) in ethanol (160 ml) wasadded 10% palladium on carbon paste (1.20 g). The mixture washydrogenated at 20 psi overnight at room temperature. Filtration throughkieselguhr and evaporation in vacuo gave the title compound as a darkgum (0.795 g, 96%). Mass spectrum (API⁺): 251 (MH⁺). C₁₃H₁₈ N₂O₃requires 250.

[0091] Description 5:(RS)-4-Benzyl-3-(t-butyloxycarbonylaminomethyl)morpholine

[0092] A solution of di-t-butyl dicarbonate (1.20 g, 5.49 mmol) in MDC(5 ml) was added dropwise, with ice cooling, to a stirred solution of D2(1.13 g, 5.49 mmol) in MDC (30 ml). The resulting solution was stirredovernight at room temperature, washed with saturated aqueous sodiumhydrogen carbonate and the organic layer evaporated in vacuo. Theresidue was purified on silica gel eluting with ethyl acetate/hexanemixtures to give the title compound as a colourless gum (1.68 g, 100%).Mass spectrum (API⁺): 307 (MH⁺). C₁₇H₂₆N₂O₃ requires 306.

[0093] Description 6: (RS)-3-(t-Butyloxycarbonylaminomethyl)morpholine

[0094] To a solution of D5 (1.68 g, 5.82 mmol) in ethanol (200 ml) wasadded 10% palladium on carbon paste (1.80 g). The mixture washydrogenated at 20 psi overnight at room temperature. Filtration throughkieselguhr and evaporation in vacuo gave the title compound as acolourless oil (1.16 g, 92%). Mass spectrum (API⁺): Found 217 (MH⁺).C₁₀H₂₀N₂O₃ requires 216.

[0095] Description 7:(RS)-3-(t-Butyloxycarbonylaminomethyl)-4-((4-(2-methyl-5-(4-fluorophenyl))thiazolyl)carbonyl)morpholine

[0096] 2-Methyl-5-(4-fluorophenyl)thiazole-4-carbonyl chloride (1.73 g,6.75 mmol) in MDC (20 ml) was added to a solution of D6 (1.16 g, 5.37mmol) and triethylamine (2.24 ml, 16.1 mmol) in MDC (80 ml) and themixture stirred at ambient temperature overnight. The reaction mixturewas washed with saturated aqueous sodium hydrogen carbonate (50 ml). Theorganic layer was evaporated in vacuo and the resultant residuechromatographed on silica gel eluting with ethyl acetate/hexane mixturesto give the title compound as a golden oil (0.73 g, 31%). Mass spectrum(API⁺): 436 (MH⁺). C₂₁H₂₆FN₃O₄S requires 435.

[0097] Description 8:(RS)-3-(Aminomethyl)-4-((4-(2-methyl-5-(4-fluorophenyl))thiazolyl)-carbonyl)morpholine

[0098] A solution of D7 (730 mg, 1.68 mmol) in MDC (15 ml) andtrifluoroacetic acid (1.5 ml) was stirred at 40° C. for 0.5 h. Thesolution was evaporated, and the resulting oil dissolved in 0.5M HCl (20ml) and washed twice with ethyl acetate (20 ml). The aqueous phase wasbasified to pH 14 with aqueous NaOH, then extracted with MDC (3×50 ml).The combined organic extracts were dried (Na₂SO₄) and evaporated to acolourless gum (462 mg, 82%). Mass spectrum (API⁺): 336 (MH⁺).C₁₆H₁₈FN₃O₂ S requires 335.

EXAMPLE 1

[0099](RS)-3-(2-Methoxybenzamidomethyl)-4-((4-(2-methyl-5-phenyl))thiazolyl)carbonyl)morpholine

[0100] A mixture of D4 (30 mg, 0.12 mmol),2-methyl-5-phenylthiazole-4-carbonyl chloride (31 mg, 0.13 mmol), andtriethylamine (36 mg, 0.35 mmol) in MDC (5 ml) was shaken for 1 h. Theresultant solution was washed with saturated aqueous sodium hydrogencarbonate (6 ml). The organic layer was applied directly onto a dry 10 gpre-packed silica cartridge and eluted with 30-100% ethyl acetate inhexane to give the title compound as an off white solid (32 mg, 60%).Mass spectrum Electrospray LC MS): Found 452 (MH⁺). C₂₄H₂₅N₃O₄S requires451.

EXAMPLE 2

[0101](RS)-3-((4-Benzofuranyl)carbonylaminomethyl)-4-((4-(2-methyl-5-(4-fluoro-phenyl))thiszolyl)carbonyl)morpholine

[0102] A mixture of D8 (30 mg, 0.09 mmol), benzofuran-4-carboxylic acid(20 mg, 0.12 mmol), 1-(3-dimethylaminopropyl)-3-ethyl carbodiimidehydrochloride (17 mg, 0.09 mmol) and 1-hydroxybenzotriazole (10 mg) inMDC (5 ml) was shaken for 20 h. The resulting solution was washed withsaturated aqueous sodium hydrogen carbonate (6 ml). The organic layerwas applied directly onto a dry 10 g pre-packed silica cartridge andeluted with 30-100% ethyl acetate in hexane, then 2-20% methanol inethyl acetate to give the title compound as an off-white solid (31 mg,72%). Mass spectrum (Electrospray LC MS): Found 480 (MH⁺). C₂₅H₂₂FN₃O₄Srequires 479.

[0103] The compounds of the examples below were prepared from theappropriate amine and acid using similar procedures to those describedabove.

Mass Spectrum Example R¹ R² (Electrospray LC/MS)  3*

Found 480 (MH⁺) C₂₅H₂₂FN₃O₄S requires 479.  4

Found 479 (MH⁺) C₂₅H₂₃FN₄O₃S requires 478.  5

Found 498 (MH⁺) C₂₅H₂₄FN₃O₅S requires 497.  6

Found 495 (MH⁺) C₂₅H₂₃FN₄O₄S requires 494.  7

Found 524 (MH⁺) C₂₄H₂₁F₄N₃O₄S requires 523.  8

Found 491 (MH⁺) C₂₆H₂₃FN₄O₃S requires 490  9

Found 463 (MH⁺) C₂₄H₂₂N₄O₄S requires 462 10

Found 480 (MH⁺) C₂₅H₂₂FN₃O₄S requires 479 11

Found 480 (MH⁺) C₂₅H₂₂FN₃O₄S requires 479 12

Found 448 (MH⁺) C₂₄H₂₁N₃O₄S requires 447

EXAMPLE 13

[0104](RS)-3-(2-Methoxybenzamidomethyl)-4-((4-(2-methyl-5-(4-fluorophenyl))thiazolyl)carbonyl)morpholine

[0105] A mixture of D4 (50 mg, 0.20 mmol),2-methyl-5-(4-fluorophenyl)thiazole-4-carboxylic acid (52 mg, 0.22mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (38mg, 0.20 mmol) and 1-hydroxybenzotriazole (10 mg) in MDC (5 ml) wasshaken for 20 h. The resultant solution was washed with saturatedaqueous sodium hydrogen carbonate (6 ml). The organic layer was applieddirectly onto a dry 10 g pre-packed silica cartridge and eluted with30-100% ethyl acetate in hexane to give the title compound as an offwhite solid (53 mg, 57%). Mass spectrum (Electrospray LC MS): Found 470(MH⁺). C₂₄H₂₄FN₃O₄S requires 469.

[0106] It is to be understood that the present invention covers allcombinations of particular and preferred subgroups described hereinabove.

[0107] Determination of Orexin-1 Receptor Antagonist Activity

[0108] The orexin-1 receptor antagonist activity of the compounds offormula (I) was determined in accordance with the following experimentalmethod.

[0109] Experimental Method

[0110] HEK293 cells expressing the human orexin-1 receptor were grown incell medium (MEM medium with Earl's salts) containing 2 mM L-Glutamine,0.4 mg/mL G418 Sulphate from GIBCO BRL and 10% heat inactivated fetalcalf serum from Gibco BRL. The cells were seeded at 20,000 cells/100μl/well into 96-well black clear bottom sterile plates from Costar whichhad been precoated with 10 μg/well of poly-L-lysine from SIGMA. Theseeded plates were incubated overnight at 37° C. in 5% CO₂.

[0111] Agonists were prepared as 1 mM stocks in water:DMSO (1:1). EC₅₀values (the concentration required to produce 50% maximal response) wereestimated using 11× half log unit dilutions (Biomek 2000, Beckman) inTyrode's buffer containing probenecid (10 mM HEPES with 145 mM NaCl, 10mM glucose, 2.5 mM KCl, 1.5 mM CaCl₂, 1.2 mM MgCl₂ and 2.5 mMprobenecid; pH7.4). Antagonists were prepared as 10 mM stocks in DMSO(100%). Antagonist IC₅₀ values (the concentration of compound needed toinhibit 50% of the agonist response) were determined against 3.0 nMhuman orexin-A using 11× half log unit dilutions in Tyrode's buffercontaining 10% DMSO and probenecid.

[0112] On the day of assay 50 μl of cell medium containing probenecid(Sigma) and Fluo3AM (Texas Fluorescence Laboratories) was added (Quadra,Tomtec) to each well to give final concentrations of 2.5 mM and 4 μM,respectively. The 96-well plates were incubated for 90 min at 37° C. in5% CO₂. The loading solution containing dye was then aspirated and cellswere washed with 4×150 μl Tyrode's buffer containing probenecid and 0.1%gelatin (Denley Cell Wash). The volume of buffer left in each well was125 μl. Antagonist or buffer (25 μl) was added (Quadra) the cell platesgently shaken and incubated at 37° C. in 5% CO₂ for 30 min. Cell plateswere then transferred to the Fluorescent Imaging Plate Reader (FLIPR,Molecular Devices) instrument and maintained at 37° C. in humidifiedair. Prior to drug addition a single image of the cell plate was taken(signal test), to evaluate dye loading consistency. The run protocolused 60 images taken at 1 second intervals followed by a further 24images at 5 second intervals. Agonists were added (by the FLIPR) after20 sec (during continuous reading). From each well, peak fluorescencewas determined over the whole assay period and the mean of readings 1-19inclusive was subtracted from this figure. The peak increase influorescence was plotted against compound concentration and iterativelycurve fitted using a four parameter logistic fit (as described by Bowenand Jerman, TiPS, 1995, 16, 413-417) to generate a concentration effectvalue. Antagonist Kb values were calculated using the equation:

K _(b) =IC ₅₀/(1+([3/EC ₅₀])

[0113] where EC₅₀ was the potency of human orexin-A determined in theassay (in nM terms) and IC₅₀ is expressed in molar terms.

[0114] Compounds of Examples tested according to this method had pKbvalues≧6.8 at the human cloned orexin-1 receptor.

[0115] The orexin-2 receptor antagonist activity of the compounds offormula (I) was determined in accordance with the following experimentalmethod.

[0116] Experimental Method

[0117] CHO-DG44 cells expressing the human orexin-2 receptor were grownin cell medium (MEM medium with Earl's salts) containing 2 mML-Glutamine, 0.4 mg/ML G418 Sulphate from GIBCO BRL and 10% heatinactivated fetal calf serum from Gibco BRL. The cells were seeded at20,000 cells/100 μl/well into 96-well black clear bottom sterile platesfrom Costar which had been pre-coated with 10 μg/well of poly-L-lysinefrom SIGMA. he seeded plates were incubated overnight at 37° C. in 5%CO₂.

[0118] Agonists were prepared as 1 mM stocks in water:DMSO (1:1). EC₅₀values (the concentration required to produce 50% maximal response) wereestimated using 11× half log unit dilutions (Biomek 2000, Beckman) inTyrode's buffer containing probenecid (10 mM HEPES with 145 mM NaCl, 10mM glucose, 2.5 mM KCl, 1.5 mM CaCl₂, 1.2 mM MgCl₂ and 2.5 mMprobenecid; pH7.4). Antagonists were prepared as 10 mM stocks in DMSO(100%). Antagonist IC₅₀ values (the concentration of compound needed toinhibit 50% of the agonist response) were determined against 10.0 nMhuman orexin-A using 11× half log unit dilutions in Tyrode's buffercontaining 10% DMSO and probenecid.

[0119] On the day of assay 50 μl of cell medium containing probenecid(Sigma) and Fluo3AM (Texas Fluorescence Laboratories) was added (Quadra,Tomtec) to each well to give final concentrations of 2.5 mM and 4 μM,respectively. The 96-well plates were incubated for 60 min at 37° C. in5% CO₂. The loading solution containing dye was then aspirated and cellswere washed with 4×150 μl Tyrode's buffer containing probenecid and 0.1%gelatin (Denley Cell Wash). The volume of buffer left in each well was125 μl. Antagonist or buffer (25 μl) was added (Quadra) the cell platesgently shaken and incubated at 37° C. in 5% CO₂ for 30 min. Cell plateswere then transferred to the Fluorescent Imaging Plate Reader (FLIPR,Molecular Devices) instrument. Prior to drug addition a single image ofthe cell plate was taken (signal test), to evaluate dye loadingconsistency. The run protocol used 60 images taken at 1 second intervalsfollowed by a further 24 images at 5 second intervals. Agonists wereadded (by the FLIPR) after 20 sec (during continuous reading). From eachwell, peak fluorescence was determined over the whole assay period andthe mean of readings 1-19 inclusive was subtracted from this figure. Thepeak increase in fluorescence was plotted against compound concentrationand iteratively curve fitted using a four parameter logistic fit (asdescribed by Bowen and Jerman, TiPS, 1995, 16, 413-417) to generate aconcentration effect value. Antagonist Kb values were calculated usingthe equation:

Kb=IC50/(1+([3/EC50])

[0120] where EC50 was the potency of human orexin-A determined in theassay (in nM terms) and IC50 is expressed in molar terms.

[0121] Compounds of Examples tested according to this method had pKbvalues in the range 6.1-7.4. at the human cloned orexin-2 receptor.

[0122] The application of which this description and claims forms partmay be used as a basis for priority in respect of any subsequentapplication. The claims of such subsequent application may be directedto any feature or combination of features described herein. They maytake the form of product, composition, process, or use claims and mayinclude, by way of example and without limitation the following claims:

1 1 1 35 PRT Homo Sapien 1 Glu Pro Leu Pro Asp Cys Cys Arg Gln Lys ThrCys Ser Cys Arg Leu 1 5 10 15 Tyr Glu Leu Leu His Gly Ala Gly Asn HisAla Ala Gly Ile Leu Thr 20 25 30 Leu Asn His 35

1. A compound of formula (I):

wherein: R¹ is phenyl, naphthyl, a mono or bicyclic heteroaryl groupcontaining up to 3 heteroatoms selected from N, O and S; any of whichmay be optionally substituted; R² represents phenyl or a 5- or6-membered heteroaryl group containing up to 3 heteroatoms selected fromN, O and S, wherein the phenyl or heteroaryl group is substituted by R³,and further optional substituents; or R² represents an optionallysubstituted bicyclic aromatic or bicyclic heteroaromatic groupcontaining up to 3 heteroatoms selected from N, O and S; R³ representsan optionally substituted (C₁₋₄)alkoxy, halo, optionally substituted(C₁₋₆)alkyl, optionally substituted phenyl, or an optionally substituted5- or 6-membered heterocyclic ring containing up to 3 heteroatomsselected from N, O and S; or a pharmaceutically acceptable salt thereof.2. A compound according to claim 1 wherein R¹ is an optionallysubstituted phenyl, benzofuranyl, quinolinyl, indolyl or benzoxazolyl.3. A compound according to claim 1 or 2 wherein R² represents anoptionally substituted thiazolyl.
 4. A compound according to any one ofclaims 1 to 3 wherein R³ represents an optionally substituted phenyl orpyridyl group.
 5. The compound of any one of Examples 1 to 13 or apharmaceutically acceptable salt of any one thereof.
 6. A pharmaceuticalcomposition comprising a compound of formula (I) as defined in any oneof claims 1 to 5, or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier.
 7. A method of treating orpreventing diseases or disorders where an antagonist of a human orexinreceptor is required, which comprises administering to a subject in needthereof an effective amount of a compound of formula (I) as defined inany one of claims 1 to 5, or a pharmaceutically acceptable salt thereof.